Mycobacterium tuberculosis (Mtb) infects about one third of the population worldwide. An estimated 8 million new cases and 2 million deaths occur annually placing tuberculosis (TB) as a significant health problem. The current vaccine for TB is ineffective in preventing transmission of TB. Furthermore, an increasing number of US military personnel are being deployed to TB endemic regions such as Afghanistan. There is an urgent need to develop of an efficacious vaccine for Mtb that will require a better understanding of the cellular immune response that is critical to control the disease. While the adaptive cellular immune response is essential in the host control of TB, many of the innate immune mechanisms that are required to result in an optimal adaptive cellular immune response to Mtb remain to be characterized. Mtb is contracted by aerosol exposure and the majority of TB cases are exclusively pulmonary. Nonetheless, little is known about the early events that occur after exposure to Mtb in the human airway and lung and basic characterization of the initial infection in humans is lacking. Upon exposure to Mtb, the epithelial cells that line the airways are the cells most likely to encounter the aerosolized bacteria. We recently determined that primary human large airway epithelial cells once infected with Mtb can serve as targets for an innate T cell population known as mucosal associated invariant T (MAIT) cells that preferentially localizes to the lung. The long-term goal of this research is to identify the innate cellular effectors that play a role in the early immune response to Mtb in the human airway and lung. In Aim 1 we want to define the cells in the human airway that can be infected with Mtb and then determine which of those cells can activate innate MAIT cells. AIM 1. Identify the airway cells that are infected with Mtb and those that activate innate MAIT cells. Hypothesis: Innate MAIT cells are general detectors of Mtb-infected cells SA1a. Use immunofluorescence and confocal microscopy to identify where Mtb localizes in airway tissue SA1b. Identify airway cells that can be infected with Mtb and activate innate MAIT cells Epithelial cells at mucosal sites provide a first line of defense against foreign pathogens. Airway dendritic cells (DC), an essential element in priming subsequent adaptive immunity, are located above and below the basement member of the epithelium. In infection with Mtb, DC are essential in priming the Mtb-specific adaptive T cells that are crucial to control the pathogen. In this proposal we want to address the key question of how DC acquire Mtb antigens in the airway and if innate T cells can enhance antigen presentation. AIM 2. Determine how airway dendritic cells (DC) acquire Mtb antigens. Hypothesis: Dendritic cells acquire Mtb antigens from Mtb-infected airway epithelial cells. SA2a. Assess how DC acquire Mtb antigens from epithelial cells SA2b. Determine the role of MAIT cells in antigen presentation by DC We have shown that MAIT cells make up a demonstrable proportion of the Mtb-reactive nonclassical CD8+ T cell response in humans. However, our work suggests that additional nonclassical CD8+ T cell populations remain to be characterized. In Aim 3 we propose to clone and characterize these nonclassical Mtb-reactive CD8+ T cells to further define the innate T cell response to Mtb in humans. AIM 3. Characterize undefined subsets of nonclassically restricted Mtb-reactive T cells. Hypothesis: Nonclassical non-MAIT Mtb-reactive T cells represent novel innate T cell subsets. SA3a. Clone and characterize non-MAIT CD8+ 34 TCR- T cells that are Mtb-reactive